Both cortical thymocytes and "mature" medullary thymocytes are derived from dividing lymphoblastoid precursors in the thymus. It is unknown whether the precursor cells are already committed to helper or cytotoxic lineages, and whether they themselves are immunocompetent. We will isolate these stem cells from the thymus and test them directly for functional reactivity. We will subfractionate them according to their expression of peanut agglutinin binding sistes, thymus leukemia antigen, and the lineage marker Lyt-2/3, and measure the stability of these phenotypic characteristicss in vitro. The functional reactivities of the different blast cell populations will then be compared, using limiting dilution cultures to determine the frequencies of responding cells. The assays will measure colony formation in response to alloantigens or polyclonal mitogens, and inducible secretion of the helper factor interleukin-2. We will also test whether the pre-killer thymic lymphoblasts exhibit as strong a preference for class I histocompatibility antigen targets as pre-killer cells that have emigrated from the thymus, and whether the early pre-helper cells already show a preference for class II targets. To dissect the development of immune competence, we will study the regulation of IL-2 receptor expression in stem cells, postmitotic thymocytes, and peripheral T cells: thus, we will test whether populations that mount weak proliferative responses are defective in their expression of this receptor. Finally, we will lay the groundwork for a molecular analysis of T-cell triggering throughout development by using DNA-mediated gene transfer and molecular cloning to isolate the gene for the mouse IL-2 receptor.